The mechanism by which dopamine inhibits prolactin secretion in the rat will be investigated in four major objectives: 1) To correlate between dopamine and prolactin inhibiting factor (PIF) activity in portal blood and the posterior pituitary. The levels of dopamine in portal blood and prolactin in the systems blood will be correlated during acute alterations in prolactin secretion caused by pup suckling. PIF activity in blood and tissue extracts will be determined by isolated rat anterior pituitary cells utilizing specific dopamine antagonists and extraction procedures for removal of catecholamines. 2) To study the role of the posterior pituitary in regulating prolactin secretion. The effects of posterior pituitary lobectomy in male and female rats on prolactin, LH and GH secretions will be studied. The dopaminergic system of the posterior lobe will be characterized in terms of the origin of dopamine, its in situ synthesis, and possible changes in its concentration and/or turnover under various endocrine states. 3) To characterize dopaminergic receptors in anterior pituitary cells. Binding parameters such as affinity and receptor density will be determined in mixed populations of isolated cells, and in fractions enriched with lactotrophs after cell separation by density gradients. Correlation between binding parameters and different endocrine states and the metabolic and ionic requirements for binding will also be studied. 4) To investigate the role of calcium and membrane potential in the inhibition of prolactin secretion by dopamine. Membrane potential and changes in free intracellular calcium will be determined by conventional and calcium-selective microelectrodes before and after exposure of the cells to inhibitors and stimulators of prolactin secretion. The potential strength of this proposal lies in the direct correlation between secretory events at the cellular and membrane level and systematic studies of pituitary secretion in the whole animal.